Process of producing acetone and butyl alcohol by fermentation



Patented Jan. 10, 1928.

UNITED STATES EDWIN F. PIKE, OF CHESTER, AND HENRY F. SMY'IH, OF W'AYNE, PENNSYLVANIA, AS-

PATENT OFFICE.

SIGNORS, BY MESNE ASSIGNMENTS, T0 COMMERCIAL SOLVENTS CORPORATION, A

CORPORATION OF MARYI AN D.

PROCESS OF PRODUCING ACETONE AND BUTYL ALCOHOL BY FERMENTATION.

No Drawing.

Our invention relates to a process of producing acetone and butylalcohol by fer- Ulostridz'um butym'cmn, the acetone and butyl alcohol,after the process of fermentation is complete, being separated from thefermented mass by suitable distillation as hereinafter described.

The bacteria which we prefer to use and which have been isolated in themanner hereinafter stated, we will now proceed to describe in accordancewith the descriptive chart of the Society of American Bacteriolog'istsin order to facilitate their identification:

Source-corn mash; name-fllostridium butym'cum (Prazmowski-Pike-Smyth).

Rods .75 1 31O microns, occurring singly, in pairs and long chains.Actively motile in young cultures. Spores oval, central to excentric 122.5 microns. Cells store glycogen as granules, polar or excentric alongrod margin. Gram-positive in young cultures but early becomegram-negative. May have slender forms .5 microns in width and filamentsup to 20 microns in length.

Gelatin stab: no liquefaction.

Serum agar colonies: (anaerobic) small, fiat, irregular, grayish,semitransparent.

Agar and dextrose agar slants: (anaerobic) grayish, flat, moist,spreading, with irregular lacerate margin. No surface growthaerobically.

Dextrose broth: turbid.

' Litmus milk: acid, coagulation, abundant gas formation with firmcontraction of torn curd.

Early reduction of litmus.

Potato: (anaerobic) thin, spreading, barely visible, yellowish-whitelayer, forming later raised points. Substance of potato slant, becomingsoft and friable with gas bubbles.

Acid gas and alcohols formed in dextrose, lactose, sucrose and. starch.Acid principally butyric.

Application filed May 29, 1923. Serial No. 642,321.

Gas formed consists of CO and H Optimum temperature range 3040 C.

Colonies in agar shake cultures, compact, irregular nucleus .25.5 mm.surrounded by a fiocose haze 2-3 mm. in diameter.

Anaerobic agar stab, granular filiform growth.

Nitrates reduced to nitrites and ammonia in anaerobic culture.

No indol formation.

Isolation of the organism.

The bacillus was originally derived from corn meal by the method ofWeizmann, being obtained in predominating culture in test tubes.Thereafter, the bacillus was secured in practically pure culture byseveral successive sub-cultures in deep dextrose agar tubes, thematerial for the sub-cultures being fished from the bottom of such tubesafter cutting them open with a file and the subculture tubes beingheated to 100 C. for two minutes before incubation. After the pureculture was obtained, the bacillus has been retained in 5%10% corn mealmash.

For a specific example of the operation of our invention for thefermentation of molasses, such as blackstrap, to produce acetone andbutyl alcohol, a mash was prepared consisting of black strap dilutedwith water so as to contain substantially 5% of molasses, and was thensterilized in a suitable sterilizer by heating under approximately 15pounds steam pressure for substantially 30 minutes. The sterilizedmixture was then placed in a sterilized container and inoculated with.the culture of the bacillus. Preliminary to inoculation, a sealed sporetube containing the bacillus was opened, emptied into a flask of sterile5% corn meal, in water, and the mixture incubated at 38-42 C. for from48 to 72 hours. The contents'of this flask were then poured into themash,

- about one part of the seeding culture to 30-35 parts'of the mash beingemployed. Fermentation Wasthen allowed to proceed for substantially 4.days under temperatures,

of a sample "of the fermented mass disclosed I the presence of acetone.The mass was then steam distllled in a suitable still until thedistillate no longer smelled. of acetone or arated into two layers whichwere then separately distilled and the acetone and butyl alcoholcollected from them at their respective boiling temperatures.

It is to be noted that satisfactory results in thetreatment of the blackstrap were not obtained when the inoculated mass was contained in copperreceptacles, but that the fermentation proceeded satisfactorily when themass was contained in glass or porcelain lined receptacles, and it istherefore believed that in the commercial practice of the processthereceptacles employed should be lined v .with glass, porcelain or someother vitreous or substantially similar material.

We have further found that the addition of nltrogeneous material to themash or mixture is ordinarily unnecessary for the attainment ofsatisfactory results, the black strap molasses containing sufficientquantlties of such material to permit proper growth of the bacilli, andthe same is equally true when other carbohydrates containing suitablequantities of nitrogeneous material are being treated. Should, however,the particular material forming the base of the fermentable mixture bedeficient in nitrogeneous material, a suitable amountthereof may beadded to assist in the proper devel-' opment of the bacilli.

Moreover, while if desired, the fermenta-- tion may be carried out underanaerobic conditions, it is unnecessary for successful operation toexclude air from access to the fermenting mass and the fermentation maytherefore, and preferably, be performedunder aerobic conditions. It is,however, .de sirable to prevent access to the fermenting mass oforganisms which might be inimical to the fermentation or which might inother ways be detrimental, and we therefore take. suitable and usualprecautions to exclude such organisms as faras possible and therebyprevent the danger of resulting contamination. Thus, when the mass is,subjected to fermentation in one receptacle and to distillation inanother, the fermenting tank may be closed with a loosely fitting coveror with a tightly fitting cover provided with means to permit the escapeof the 00,, H and other products of fermentation, or when the sameapparatus is'used to hold the mash during the fermentation period andalso during the initial distilling operation, the

Same may be provided with a steam tight {cover from which a vent pipemay be extended upwardly and bent over and'down- :wardly adjacent itsextremity so as to progyvlde a long and tortuous passage, thereby verylargely preventing the entrance of bacteria to the receptacle but stillallowing the escape of theiproducts of fermentation.

It will be understood that after the initial, preferably steam,distillation, the distillate so obtained is again distilled in anysuitable performed in any suitable apparatus as will be readilyunderstood by those familiar with the act.

We are aware that it has been proposed to produce acetone and butylalcohol by the fermentation of amylaceous carbohydrates by means of anaerobic organism and that various eiforts have been made in the past toadapt such organisms to the fermentation of saccharine carbohydrates,but we believe it broadly new to produce acetone and butyl alcohol bythe fermentation ofmolasses in the absence of amylaceous materials, andwe ascribe the success of our improved process and invention to thedistinctive character of the herein described organism which producesacetone and butyl alcohol with satisfactory yields in the absence ofamylaceous materials. 7

While we have herein, for the purpose of enabling those skilled in theart to compre'-.

bend and practise the invention, described with considerableparticularity certain preferred methods of performing the same, we donot thereby desire or intend to specifically limit ourselves thereto, asvarious changes and modifications may be made for the purpose offacilitating the practice of the invention or for such other purposes asmay be desirable Without departing from the spirit and scope of theinvention as defined in the appended.- claims.

Having thus described our invention,.we claim and desire to protect byLetters Patent of the United States: a I

-l. The process of producing acetone and butyl alcohol by fermentationof molasses which comprises adding a culture 0 Glostfidium butym'cwm(Prazmowski-Pike- Smyth) in the absence of amylaceous materials to asterilized mash of molasses and water, and then allowlng fermentation toproceed to completion.

2; The process of producing acetone and butyl alcohol by fermentation ofmolasses which comprises adding a culture of Clostm'dim butyrioum(Prazmowski-Pike- Smyth) in the absence of amylaceous ma-,

5 terials to a sterilized aqueous mash containing substantially 5% ofmolasses, and then maintaining the temperature of the inoculated mash at3941 C. until fermentation is complete.

10 I 4. The process of producing acetone and butyl alcohol byfermentation of molasses which comprises preparing a sterile mash ofmolasses and water, inoculating the mash with a culture of C'Zostrid-iumbuty'riowm 16 (Prazmowski-Pike-Smyth) in the absence of amylaceousmaterials, maintaining the temperature of said inoculated mash at 3941C. until fermentation is complete, and distillin the fermented mass toobtain acetone an butyl alcohol therefrom.

5. The process of producing acetone and butyl alcohol by fermentation ofmolasses which comprises preparing a sterile mash of molasses and waterhaving a content of substantially 5% of molasses, inoculating the mashwith a culture of Glostfldium butyricum' (Prazmowski-Pike-Smyth) in theabsence of amylaceous materials, maintaining the temperature of saidinoculated mash at 3941 C. until fermentation is complete, anddistilling the fermented mass to obtain acetone and butyl alcoholtherefrom.

In witness whereof, we have hereunto set our hands this 28 day of May,1923.

r EDWIN F. PIKE.

' HENRY F. SMYTH.

